Characterization of a Novel Microtubule Binding Protein CLIPR76

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A combination of cell biological, molecular biological and biochemical analyses was used to study a novel member of ClipR family, CLIPR76. Sequence analysis and experimental evidence show that the CLIPR76 gene is alternatively spliced and produces multiple isoforms in different organisms. Western blot and real time PCR investigations show that the CLIPR76 gene is ubiquitously expressed with highest levels in testis and striated muscle. Some isoforms appear to be tissue specific. Immunofluorescence microscopy of the GFP tagged or untagged CLIPR76 proteins demonstrate that the different isoforms have different cellular localization and imply that they have different protein activities. Some CLIPR76 isoforms localize to the endoplasmic reticulum and their overexpression affect the morphology of ER and ERGIC membrane compartments, while others localize to microtubules or subsets of microtubules and cause alterations in microtubule organization when expressed at higher levels. These results imply a role for CLIPR76 proteins in membrane transport and ER microtubules interactions.and protein expression in mouse tissues and RT-PCR to investigate the
expression of CLIPR76 splice variants in the mammalian cells used in this work.
... RNA was converted to cDNA using SuperScript II Reverse transcriptase (
Invitrogen) following manufacturera#39;s instructions. ... Biosystems Prism 7700
Sequence Detection System Real-time PCR instrument or on the 7500 Fast Real-
time PCR System.

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Characterization of a Novel Microtubule Binding Protein CLIPR76

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ProQuest - 2007

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